Antifungal parenteral products

ABSTRACT

Parenteral pharmaceutical formulations containing an echinocandin antifungal compound and an aqueous solvent are provided, wherein the formulation includes ethanol, for example about 20% w/v ethanol. The parenteral pharmaceutical formulation may further include one or more additives, such as a stabilizing agent, buffer or tonicity agent. The parenteral pharmaceutical formulations are useful in extending the shelf life and improving the solubility of the echinocandin antifungal compound.

TECHNICAL FIELD

[0001] This invention relates to drug formulations, for example, aqueousinjectable drug formulations and methods for their manufacture and use.

BACKGROUND OF THE INVENTION

[0002] One aspect of the commercial viability of an injectable drugproduct is long shelf life. A shelf life significantly greater than oneyear is typically needed. This is because drug products are often storedfor long periods, for example, six months to a year or more, untilneeded. The expiration date of a product begins when the drug isproduced, but testing and packaging for shipping often take up sometime, for example, months. A shelf life of one to three years or more isvery desirable for an injectable drug product. This is especially truefor a drug which may be stored for a long period of time, because it isnot frequently used, but that is specifically required when indicated.

[0003] Another aspect of injectable drug products is reconstitution ofthe formulation by the medical practitioner. The drug may be deliveredin a solid form, often called “drug for injection,” which may containother ingredients, and is reconstituted to a liquid form by the additionof solvent and other components.

[0004] A USP (United States Pharmacopeia) requirement for parenteraldrug products is that the product be visibly clear before use. A vial ofcrystal clear liquid is desired. To meet this standard, the number ofparticulates in the reconstituted liquid product must be kept to aminimum. Particulates represent undissolved drug which is ineffective,and may block capillaries causing serious adverse health effects. Acrystal clear drug product requires the solution to have a minimum ofminute, undissolved, or non-visible drug particles.

[0005] Particulates in an injectable drug product may be caused in partby foaming during reconstitution. Foaming may be caused by the drugitself, or by surfactants other additives used to increase and hastensolubility of the drug. Foaming can prevent small particles fromentering the solution to be dissolved, thereby increasing the number ofparticulates in the reconstituted injectable drug product.

[0006] Another aspect of parenteral drug products is the time requiredto reconstitute an injectable formulation. Reconstitution requires theability to rapidly redissolve a drug composition to provide a crystalclear solution. In addition, the reconstitution should be rapid after along storage period of the delivered drug composition, a period whichcan be, for example, one, two, or three years or more. Some drugcompositions as delivered typically cannot achieve a shelf life greaterthan one year. This is because either the reconstitution time is toolong after storage, or the number of particulates in the reconstitutedproduct is too high.

[0007] Echinocandin antifungal compounds, and methods for theirmanufacture and use are described, for example, in PCT WO 00/52037; PCTWO 00/51564; PCT WO 00/34315; PCT WO 00/51567; and U.S. Pat. No.5,965,525.

[0008] There is a need for echinocandin pharmaceutical drug formulationsthat are useful for parenteral pharmaceutical administration with rapidreconstitution, little forming, and a long shelf life.

SUMMARY OF THE INVENTION

[0009] In one aspect, a pharmaceutically acceptable parenteralformulations containing an echinocandin and an aqueous ethanolic solventis provided, as well as optionally one or more additives, such aspropylene glycol, or polyethylene glycol, a buffer, stabilizing agent,tonicity agent, antioxidant or bulking agent. Methods of reconstitutingsolid compositions containing an echinocandin also are provided.

[0010] In one embodiment, pharmaceutically acceptable liquidformulations are provided that include, e.g., about 0.2 to 1.0%, orabout 0.1 to 2.0% or about 0.2 to 2.0% w/v anidulafungin. Thepharmaceutically acceptable liquid formulations also may include, e.g.,about 1.0 to 4.0 mg/mL, or about 1.5 to 3.0 mg/mL anidulafungin.

[0011] A pharmaceutically acceptable liquid parenteral formulation forexample is provided comprising: anidulafungin and an aqueous solventsuch as water or saline, wherein the formulation includes from about 5%w/v ethanol to about 50% w/v ethanol; about 15-30% w/v ethanol; or about20% w/v ethanol. The formulation may include about 10% to about 40% w/vethanol and about 0.2 to about 2.0% w/v anidulafungin, or optionallyabout 15 to about 30% w/v ethanol or optionally 20 percent w/v ethanol.The formulation also may include 10 to about 50 percent w/v propyleneglycol or polyethylene glycol, or mixtures thereof.

[0012] The formulation further may include a stabilizing agent, such asmannitol, histidine, lysine, glycine, sucrose, fructose, trehalose,lactose or a mixture thereof. The formulation can optionally contain abulking agent, such as mannitol, glycine, lactose, sucrose, trehalose,dextran, hydroxyethyl starch, ficoll or gelatin. The formulation canoptionally contain a solubilizing agent or surfactant, such ascetrimide, docusate sodium, glyceryl monooleate, sodium lauryl sulfate,or sorbitan esters. The solubilizing agent or surfactant may optionallybe a polyoxyethylenesorbitan fatty acid ester. Polyoxyethylenesorbitanfatty acid esters are also referred to as polysorbates, e.g.,polysorbate 80 (polyoxyethylene sorbitan monooleate, Tween 80),polysorbate 40 and polysorbate 20. Polysorbates, such as polysorbate 80,are commercially available, for example, from Sigma, St. Louis, Mo.

[0013] The formulation can optionally comprise a buffer, such asacetates, citrates, tartrates, lactates, succinates, or phosphates. Theformulation can optionally contain a tonicity agent, such as glycerin,lactose, mannitol, dextrose, sodium chloride, sodium sulfate orsorbitol. The formulation can optionally contain an antioxidant, such asacetone, sodium bisulfite, bisulfite sodium, butylated hydroxy anisole,butylated hydroxy toluene, cysteine, cysteinate HCl, dithionite sodium,gentisic acid, gentisic acid ethanolamine, glutamate monosodium,formaldehyde sulfoxylate sodium, metabisulfite potassium, metabisulfitesodium, monothioglycerol, propyl gallate, sulfite sodium, thioglycolatesodium, or ascorbic acid.

[0014] A pharmaceutically acceptable parenteral formulation is alsoprovided for example comprising: anidulafungin and an aqueous solvent,such as water or saline, wherein the formulation includes:

[0015] about 5-30% w/v ethanol;

[0016] about 0.1-2.0% w/v anidulafungin;

[0017] about 0.1-1.0% w/v of a stabilizing agent, such as fructose;

[0018] about 0.1-10% w/v of a bulking agent, such as mannitol;

[0019] about 0.01-5% w/v of a buffer, such as tartaric acid; and

[0020] about 0.1-5.0% w/v of a solubilizing agent, such as polysorbate80.

[0021] The formulation may optionally include about 2-50% w/vpolyethylene glycol and/or propylene glycol.

[0022] A pharmaceutically acceptable parenteral formulation is alsoprovided comprising: anidulafungin and an aqueous solvent, such as wateror saline and ethanol, wherein the anidulafungin is stored in solid formfor greater than 6, 9, 12, 15 or 18 months prior to forming theformulation, and wherein the formulation is suitable for use as aparenteral formulation.

[0023] A formulation is “suitable for use as a parenteral formulation”if it is in a pharmaceutically acceptable form for parenteraladministration. Thus, for example, for a liquid parenteral formulation,the particle content is sufficiently low, and the material issufficiently sterile such that it is useful for parenteraladministration. To be suitable for parenteral administration, theformulation is visibly clear, and the number of particles in thereconstituted liquid product is kept to a minimum. For example, lessthan 6,000 10 μm particles should be present in a volume of 10 mLsolvent that includes 35 mg of anidulafungin. For example, when the drugis freeze dried and stored, for example, for 9 months, 12 months, 18months or 24 months, and then reconstituted by dissolving 35 mg of drugand optionally other additives in 10 ml of aqueous ethanolic solvent,there are preferably less than 10,000, less than 6,000, less than 3,000,less than 1,000, or less than 400 10 μm particles. There are, forexample, less than 1000, less than 600, or less than 200 25 μm particlesin the 10 mL volume.

[0024] Liquid parenteral formulations formed from drug stored over longtime periods can become no longer suitable for parenteraladministration, because of particles in the reconstituted formulations.Using the methods disclosed herein, liquid formulations are formed whichare suitable for parenteral administration, even when formed from drugstored over long time periods. This can extend the shelf life of thedrug. Moreover, using the ethanolic solutions for reconstitution resultsin more rapid dissolution of the drug.

[0025] Also provided is a method of preparing a parenteralpharmaceutical formulation, comprising combining a solid compositioncomprising anidulafungin and an ethanolic aqueous solvent, such aswater, to substantially dissolve the anidulafungin and produce anaqueous formulation including about 5 to about 50% w/v ethanol,optionally 10 to about 30 percent w/v ethanol, wherein the formulationis optionally shaken until the mixture is substantially clear. Themethod optionally comprises forming the solid composition containinganidulafungin by lyophilizing an aqueous solution of anidulafungin andoptionally an additive such as polyethylene glycol or propylene glycol.The solid composition can optionally contain a stabilizing agent, abuffer, a tonicity agent or an antioxidant.

[0026] The method optionally comprises preparing a solid compositioncomprising anidulafungin produced for pharmaceutical use and stored formore than one year before combining with an ethanol water solvent, andwherein the combining step produces a formulation suitable for use as apharmaceutically acceptable parenteral formulation.

[0027] Also provided here is a kit for use in delivery of apharmaceutically acceptable parenteral pharmaceutical formulation,comprising: a vial containing a pharmaceutically acceptable solidformulation comprising anidulafungin; and a vial comprising apharmaceutically acceptable aqueous solution of about 10 to 30% w/vethanol. The kit can optionally contain an additive, such aspolyethylene glycol, propylene glycol, a stabilizing agent, a buffer, atonicity agent and/or an antioxidant. The kit can optionally contain avial containing a solid formulation comprising 25 to 200 mg ofanidulafungin and a second vial containing 5 to 60 milliliters ofaqueous solution comprising about 10 to 30% w/v ethanol.

[0028] Also provided are pharmaceutically acceptable parenteralformulations including anidulafungin and an aqueous solvent, wherein theanidulafungin is stored for greater than 6, 9, 12, 15, or 18 monthsbefore formation of the liquid formulation, and wherein the formulationis suitable for administration as a parenteral formulation.

[0029] Further provided are methods of preparing a parenteralformulation, comprising combining a solid formulation comprisinganidulafungin with a solvent in an effective amount to dissolve theanidulafungin rapidly, for example in 400 seconds or less, in 200seconds or less, in 100 seconds or less, or in 60 seconds or less, forexample by shaking or swirling, to produce a pharmaceutically acceptableparenteral formulation. The anidulafungin may be a formulation producedfor pharmaceutical use and stored, for example for more than one year,or two years, prior to combining with the solvent. The concentration ofthe anidulafungin is, for example, about 1.5 to 3 mg/mL, or about 1.5 to5 mg/mL.

DETAILED DESCRIPTION OF THE INVENTION

[0030] Provided are pharmaceutically acceptable formulations comprisingan echinocandin, such as anidulafungin.

[0031] Echinocandins

[0032] Echinocandin-type compounds have been shown to exhibit antifungaland antiparasitic activity, such as inhibition of growth of variousinfectious fungi including Candida spp. (i.e., C Albicans, CParapsilosis, C Krusei, C Glabrata, C Tropicalis, or C Lusitaniaw);Torulopus spp. (i.e., T Glabrata); Aspergillus spp. (i.e., A.Fumigalus), Histoplasma spp. (i.e., H. Capsulatum); Cryptococcus spp.(i.e., C. Neoformans); Blastomyces spp. (i.e., B. Dermatilidis);Fusarium spp.; Trichophyton spp., Pseudallescheria boydii, Coccidioidesimmits, and Sporothrix schenckii, etc. PCT WO 00/51564.

[0033] Compounds of this type also have been shown to inhibit the growthof certain organisms primarily responsible for opportunistic infectionsin immunosuppressed individuals, such as growth inhibition ofPneumocystis carinii. Other protozoans that are inhibited byechinocandin-type compounds include Plasmodium spp., Leishmania spp.,Trypanosoma spp., Cryptosporidium spp., Isospora spp., Cyclospora spp.,Trichomnas spp., and Microsporidiosis spp., etc. PCT WO 00/51564.

[0034] Consequently, the formulations of the present invention areuseful in the treatment of, e.g., systemic fungal infections or fungalskin infections. Accordingly, the processes and formulations of thepresent invention may be used in the manufacture of a medicament for thetherapeutic applications described herein. For example, fungal activity(preferably, Candida albicans or Aspergillus fumigatis activity) orparasitic activity may be inhibited by contacting a pharmaceuticalformulation prepared by the present invention with a fungus or parasite,respectively. The term “contacting” includes a union or junction, orapparent touching or mutual tangency of a compound of the invention witha parasite or fungus. The term does not imply any further limitations tothe process, such as by mechanism of inhibition. The methods are definedto encompass the inhibition of parasitic and fungal activity by theaction of the compounds and their inherent antiparasitic and antifungalproperties.

[0035] A method for treating a fungal infection which comprisesadministering an effective amount of a pharmaceutical formulation of thepresent invention to a host in need of such treatment is also provided.The method includes treating a Candida albicans or Aspergillus fumigatisinfection. The term “effective amount” refers to an amount of activecompound which is capable of inhibiting fungal activity. The doseadministered will vary depending on such factors as the nature andseverity of the infection, the age and general health of the host andthe tolerance of the host to the antifungal agent. The particular doseregimen likewise may vary according to these factors. The drug may begiven in a single daily dose or in multiple doses during the day. Theregimen may last for example, for 2-3 days, for 14-30 days, or longer.An exemplary daily dose (administered in single or divided doses)contains a dosage level between about 0.01 mg/kg to 100 mg/kg of bodyweight of an active compound. Further exemplary daily doses are about0.1 mg/kg to 60 mg/kg or about 0.1 mg/kg to 40 mg/kg, or about 0.7 to 3mg/kg per day. Further exemplary daily doses are about 5 to 500 mg/dayor about 50 to 200 mg per day.

[0036] Anidulafungin(1-[(4R,5R)-4,5-Dihydroxy-N(2)-[[4″-(pentyloxy)[1,1′:4′,1″-terphenyl]-4-yl]carbonyl]-L-omithine]echinocandinB) is an echinocandin that can be semisynthetically derived from anatural product. The synthesis of anidulafungin is described in U.S.Pat. No. 5,965,525.

[0037] Like other echinocandins, anidulafungin has a low watersolubility of less than 0.1 mg/ml. Because of the low solubility,formulations have been described that add a surfactant to an aqueoussolution, however this can hinder freeze drying (WO 00/51564).

[0038] It has been found in the past to be difficult to prepare aformulation of anidulafungin in water which meets the strict USPrequirements for purity and clarity of injectable formulations, evenwhen such concentrations are well below the solubility limit in thepresence of surfactant species.

[0039] The structure of anidulafungin is provided below

[0040] Solid Compositions

[0041] Solid pharmaceutical compositions containing an echinocandin areprovided that can be formulated for administration to a patient in needthereof. Such solid compositions may be reconstituted in an aqueousethanolic solvent to provide a liquid product, which may be administeredto a patient by parenteral means, including subcutaneous, intravenous,bolus injection, intramuscular, or intraarterial injection, oralternatively by oral, topical, transdermal, or mucosal administration.

[0042] Solid compositions may have crystalline and amorphous components.A solid composition of an echinocandin may be prepared by lyophilizing(freeze drying) a volume of a solution which contains a knownconcentration of the echinocandin.

[0043] A solid composition comprising anidulafungin can be formed bylyophilizing a solution of anidulafungin, such as an aqueous solution ofanidulafungin and optionally one or more additives, such as propyleneglycol, or polyethylene glycol, a buffer, stabilizing agent, tonicityagent, antioxidant or bulking agent.

[0044] The polyethylene glycol may have, for example, a molecular weightof 400 to about 1500, optionally, 600 to about 1000, often 1000. Theaddition of polyethylene glycol or propylene glycol optimizes thereconstitution of the lyophilization formulation in an aqueous solventcontaining ethanol by providing enhanced solubility. The amount ofpolyethylene glycol or propylene glycol is, for example, an amounteffective to produce the desired concentrations after addition of anaqueous solvent to form a parenteral formulation as discussed herein. Inthe solid composition, for example, a 50 mg dose of anidulafungin maycontain 1.5 grams of PEG.

[0045] Solid formulations may optionally contain a stabilizing agent.The solid formulations may contain a stabilizing reagent at aconcentration of 5 to 80% w/w, generally 7.6 to 11.5% w/w, or 9.5% w/w.The term “stabilizing agent” refers to a pharmaceutically acceptableexcipient that may enhance the chemical or physical stability of theactive ingredient in the formulation. Suitable stabilizing agentsinclude carbohydrates, such as sucrose, trehalose, fructose, andlactose, and amino acids. Other examples include polyoxyethylenesorbitanfatty esters (polysorbates), e.g., polysorbate 80 (polyoxyethylenesorbitan monooleate, Tween 80), polysorbate 40 and polysorbate 20.

[0046] Solid formulations may optionally contain a solubilizing agent orsurfactant. The solid formulations may contain a solubilizing agent at aconcentration of 10 to 50% w/w, 20 to 30% w/w, or e.g., 24% w/w.Suitable solubilizing agents include cetrimide, docusate sodium,glyceryl monooleate, sodium lauryl sulfate, and sorbitan esters.Exemplary solubilizing agents include polysorbates (e.g. polysorbate 20,polysorbate 40, polysorbate 80).

[0047] As used herein, “w/w” refers to percent weight in weight, andexpresses the number of g of a constituent in 100 g of solution ormixture.

[0048] As used herein, “w/v” refers to percent weight in volume andexpresses the number of g of a constituent in 100 mL of solution.

[0049] Solid formulations may also optionally contain a buffer. Thebuffer is optionally present at a concentration in the range from about0.3 to 5%, or about 0.9% to 1.3%, or about 1.1% w/w. The term “buffer”refers to a pharmaceutically acceptable excipient that helps to maintainthe pH of the solution within a particular range specific to thebuffering system. Suitable buffers include acetates, citrates,phosphates, tartrates, lactates, succinates, amino acids and the like.

[0050] When freeze dried, the formulations may optionally contain abulking agent. The term “bulking agent” refers to a pharmaceuticallyacceptable excipient that adds bulk to a formulation which results in awell-formed cake upon freeze drying. Suitable bulking agents includemannitol, glycine, lactose, sucrose, trehalose, dextran, hydroxyethylstarch, ficoll and gelatin. The bulking agent is for example present inthe formulation at the concentration in the range from about 30 to 68%w/w of the solid formulation. For example, the formulation comprisesabout 43 to 52% and optionally about 48% w/w bulking agent

[0051] The solid formulations may be prepared using conventionaldissolution and mixing procedures. For example, the anidulafungin isdissolved in a non-toxic aqueous solvent optionally in the presence of apharmaceutically acceptable polyethylene glycol or propylene glycol andoptionally one or more bulking agents, buffers, and/or stabilizingagents. The resulting solution is then sterilized, e.g., sterilefiltered and preferably lyophilized to provide the desired formulation.

[0052] The solution to be lyophilized may further include one or moreantioxidants, such as acetone sodium bisulfite, bisulfite sodium,butylated hydroxy anisole, butylated hydroxy toluene, cystein,cysteinate HCl, dithionite sodium, gentisic acid, gentisic acidethanolamine, glutamate monosodium, formaldehyde sulfoxylate sodium,metabisulfite potassium, metabisulfite sodium, monothioglycerol, propylgallate, sulfite sodium, thioglycolate sodium, and ascorbic acid.

[0053] The solid composition also may include a tonicity agent such asglycerin, lactose, mannitol, dextrose, sodium chloride, sodium sulfateand sorbitol.

[0054] The solution to be lyophilized may include excipients,stabilizing agents, buffers, tonicity agents, and antioxidants, asdescribed above, and may further contain agents which modify thephysical appearance and shape of the lyophilized solid such as mannitol,fructose, and tartaric acid. Solid compositions obtained fromlyophilization of the solution may include amounts of all speciesdescribed herein. Further solvents, solvent mixtures, or solvent systemsare reviewed in, S. Nema et al, PDA Journal of Pharm. Science and Tech.51(4): 166-171 (1997). The solution to be lyophilized may be sterilizedprior to lyophilization. Alternatively, the solid product fromlyophilization may be sterilized. Methods for sterilization are reviewedin Remington 's Pharmaceutical Sciences, 18th ed. (1990).

[0055] A suitable method for freeze-drying is described in Nail et al.Freeze Drying Principles and Practice, in Pharmaceutical Dosage Forms,2^(nd) Ed., Marcel Dekker, Inc. NY, pp. 163-233 (1993). The formulationis preferably freeze-dried in a vial which can then be stored untilneeded. A non-toxic, aqueous solvent is added to the vial to dissolvethe freeze-dried material thus forming a solution that can be used in aparenteral therapeutic application. Those skilled in the art willappreciate that the aqueous solvent includes other common solutions usedin such applications (e.g., saline solutions, dextrose solutions, etc.).In application, the formulations are typically diluted or reconstituted(if freeze-dried) and may be further diluted if necessary, prior toadministration.

[0056] The active ingredient is typically formulated into pharmaceuticaldosage forms to provide an easily controllable dosage of the drug and togive the patient an elegant and easily handleable product. Solidformulations may comprise for example, about 0.1% to 60% w/w of activeingredient for example, anidulafungin, or about 1% to 30% w/w, or about8.0 to 12% w/w, or optionally about 9-10% w/w.

[0057] As used herein, the term “unit dose” or “unit dosage” refers tophysically discrete units that contain a predetermined quantity ofactive ingredient calculated to produce a desired therapeutic effect.When a unit dose is administered parenterally, it is typically providedin the form of measured units in ampoules (or vials). The dosage to beadministered may vary depending upon the physical characteristics of thepatient, the severity of the patient's symptoms, and the means used toadminister the drug. The specific dose for a given patient is usuallyset by the judgment of the attending physician. Dosages can be forexample, about 5-500 mg or about 35-200 mg daily administered from anintravenous injection (IV).

[0058] Suitable carriers, diluents and excipients are known in the artand include materials such as carbohydrates, waxes, water soluble and/orswellable polymers, hydrophilic or hydrophobic materials, gelatin, oils,solvents, water, and the like. The particular carrier, diluent orexcipient used will depend upon the means and purpose for which theactive ingredient is being applied. The formulations may also includewetting agents, lubricating agents, emulsifiers, suspending agents,preservatives, sweeteners, perfuming agents, flavoring agents andcombinations thereof.

[0059] Shelf life of the solid compositions is the length of time thatthe solid composition may be stored in a form intended for a parenteralpreparation and still be reconstituted in a reasonable time. The shelflife can begin when the active drug ingredient of the solid compositionis made, and can end when the solid composition cannot be reconstitutedin a reasonable time by a particular method or when the drug degrades orotherwise cannot be used. Solid compositions with long shelf life, forexample, greater than 12 months; or greater than about 15, 20, 25, 30,35, 36 months or more are preferred. As disclosed herein, the solventadded to the solid composition to make the parenteral formulation canincrease the shelf life, to, for example, greater than 12 months; orgreater than 15, 20, 25, 30, 35, 36 months or more.

[0060] An example of solid formulations for a 35 and 50 mg dose ofanidulafungin is shown in Table A. This Table shows an exemplary amountas well as an optional range shown in parentheses of components of asolid formulation. These formulations are made in one embodiment bydissolving the ingredients in sterile water (e.g., 3-15 ml), optionallyadjusting the pH with and freeze drying. TABLE A Solid AnidulafunginFormulations Quantity in Quantity mg in mg per per vial 50 vial 35 mg mgdose dose % w/w Ingredient (mg range) (range) Function (range)Anidulafungin  50  35 Active  9.5 (40-60) (28-42) Ingredient (7.6-11.5)Fructose  50  35 Stabiliser  9.5 (45-55) (31.5-38.5) (8.6-10.5) Mannitol250 175 Bulking Agent 48 (225-275) (157-192) (43-52) Polysorbate 80 125 87.5 mg Solubilizing 24 (113-137) (79-96) Agent/ (21-26) surfactantTartaric Acid  5.6  3.94 Buffer  1.1 (5.0-6.1) (3.5-4.3) (0.9-1.3)Sodium As needed To adjust pH negligible Hydroxide Hydrochloric Asneeded To adjust pH negligible Acid

[0061] The solid or liquid formulations in a freeze dried dosage vialmay include an excess of about 1-5%, e.g., 2.5% of any one or more ofthe above components to allow for withdrawal of the required amount ofthe anidulafungin from the vial after addition of solvent and extractionfor parenteral administration.

[0062] In application, the formulations are reconstituted (iflyophilized) and are further diluted prior to administration. An exampleof reconstitution instructions for a freeze-dried product is to addaqueous ethanolic solvent to the vial and agitate to dissolve. Typicalreconstitution times are less than ten minutes, or less than one minuteThe resulting solution may be then further diluted in an infusionsolution such as dextrose 5% in water (D5W), prior to intravenousadministration.

[0063] A pharmaceutical composition may be administered using a varietyof methods. Suitable methods include injection. The particular treatmentmethod used will depend upon the type of infection being addressed.

[0064] Liquid Pharmaceutical Formulations

[0065] Provided are parenteral pharmaceutical formulations comprisinganidulafungin and an aqueous solvent, wherein the formulation includesfrom about 5% ethanol w/w to about 50 percent ethanol w/w. The aqueoussolvent, is for example, water, or e.g. saline.

[0066] As used herein, “w/w” refers to percent weight in weight, andexpresses the number of g of a constituent in 100 g of solution ormixture.

[0067] Also provided are pharmaceutically acceptable parenteralformulations comprising anidulafungin and an aqueous solvent, whereinthe formulation includes about 5% to about 50% w/w ethanol, e.g., about10 to 40%; about 15 to 30%; or about 20% w/w ethanol.

[0068] Also provided are pharmaceutically acceptable parenteralformulations comprising anidulafungin and an aqueous solvent, whereinthe formulation includes about 5 to 50% (w/w) propylene glycol and/orpolyethylene glycol (PEG) in an aqueous ethanol solution. Theseformulations may be further diluted with 5% Dextrose in Water (D5W)prior to intravenous (IV) injection.

[0069] The pharmaceutical parenteral formulations may be provided indosage vessels which contain dosage units, for example, about 5 mg to500 mg of anidulafungin, about 20 to 200 mg, or about 35 mg, 50 mg, or100 mg. The dosage vessels are often loaded with a slight excess, forexample 2.5% excess of drug, because it is generally not possible toremove all the drug from the vial after reconstitution of the drug. Forexample, a 35 mg vessel may be loaded with about 36 mg of drug. A dosageunit may be provided in a sealed container, often made of Type I glass,which maintains a sterile environment for the solid product fromlyophilization. For example, a vial hermetically sealed with a sterilerubber or plastic closure or stopper. The closure or stopper allowscharging the vial with solvent or diluent, such as sterile Water forInjection, USP, ethanol, Normal Saline, USP, or 5% Dextrose in Water,USP.

[0070] A pharmaceutically acceptable parenteral liquid productformulation may be prepared by reconstitution of a solid compositionsuch as a freeze dried solid composition, for example, as describedabove. These product liquids may be sterilized during, or afterreconstitution. The reconstitution product may be used to parenterallyadminister an echinocandin active compound, such as anidulafungin.Reconstitution may be accomplished by charging a vessel containing thesolid composition with a physiologically acceptable sterile solvent ordiluent and mixing the contents of the vessel for an acceptablereconstitution time by hand shaking or swirling. Other methods ofshaking or mixing the vessel may be used, for example, any mechanicalshaking or reciprocating mixing device. The intensity of shaking ormixing is sufficient to reconstitute the solid composition into a liquidproduct in a reasonable time, for example, within a few minutes, forexample, without causing severe foaming.

[0071] The reconstitution time of the solid composition, whenreconstituted with one of the aqueous formulations comprising ethanoldescribed herein, may be much less than the reconstitution time of thesolid composition when reconstituted with water. The reconstitution maybe carried out with an aqueous solution and ethanol as well as one ormore additives, to provide a pharmaceutically acceptable parenteralformulations.

[0072] Reconstitution time is, for example, less than 10 minutes, lessthan 5, minutes, less than 2 minutes, less than one minute, or less than45 seconds, for example after storage of a freeze dried drug formulationfor more than one year, more than two years or more than three years.Preferably, the liquid drug formulation after reconstitution includes nomore than a pharmaceutically acceptable amount of particulate matter.For example, the content of particulate matter after dissolving 35 mg ofdrug in 10 ml of aqueous ethanolic solvent, after freeze drying andstorage, for example, after one, two or three years, is less than 6000,preferably less than 400 10 μm particles, and/or less than 600,preferably less than 200 25 μm particles.

[0073] The addition of surfactant or solubilizing agent, for examplepolyoxyethylene sorbitan monooleate (Tween 80 or polysorbate 80),improves solubility but can cause foaming, which reduces exposure of thedrug to the solution and can cause less drug to be dissolved.Insolubility problems increase with increasing storage time of a freezedried material. Particles can be dispersed on top of the solution in thebubbles of the foam which makes it difficult to see in the vial if thedrug is dissolved. For example, when Tween 80 is added to increase thesolubility of anidulafungin, and the drug is freeze dried to enhancestability, the ability to redissolve the freeze dried formulation inwater becomes increasingly reduced over time. The formulations of thepresent invention overcome this disadvantage. The ethanol in thereconstitution solution can act by reducing the surface tension of thereconstituted drug formulation resulting in reduced foaming. Thisprocedure also reduces reconstitution time, due to the solubilityproperties of ethanol.

[0074] Another advantage is that reconstitution is greatly improved insolid formulations stored longer than one year. The surfactants becomeless effective over time and increased foaming is observed inreconstituted drug formulations stored 18 months or longer using waterfor reconstitution without ethanol. This may restore surfactant functionof stored solid drug formulations.

[0075] In one embodiment, the pharmaceutically acceptable parenteralformulation of echinocandin, such as anidulafungin, may have about 5 to50% w/v ethanol, e.g., ethanol USP, in water, or 20% w/v ethanol inwater or other aqueous solution. In these pharmaceutically acceptableparenteral aqueous formulations, the % w/v ethanol, such as ethanol USPmay be, for example, about 5 to about 50 percent w/v; e.g. about 10 toabout 40%; about 15 to about 30%; about 18 to 22 percent; or about 20percent w/v ethanol.

[0076] Tables B and C show examples of liquid formulations ofanidulafungin, where the formulations include, water, ethanol,anidulafungin and optionally one or more of the other components listedin the Tables; as well as exemplary ranges of the components. Theseliquid formulations may be further diluted in D5W or other aqueousdiluent for intravenous injection. TABLE B Liquid Formulations ofAnidulafungin Quantity % per vial w/v Ingredient 35 mg dose in 10 ml(range) (range) Anidulafungin   35 mg  0.34 (0.1-2.0) Fructose   35 mg 0.34 (0.1-1.0) Mannitol  175 mg  1.75 (0.1-10) Polysorbate 80 87.5 mg 0.85 (Tween 80) (0.1-5.0) Tartaric Acid 3.94 mg  0.04 (0.01-5.0) SodiumHydroxide As needed negligible Hydrochloric Acid As needed negligibleEthanol 2 g 20 (5-50) PEG and/or 1 g 10 propylene glycol (0-50) SterileWater for q.s to 10 ml to volume Injection

[0077] TABLE C Optional Ranges of Liquid Formulations of Anidulafungin %wgt./wgt. Ingredient range Anidulafungin 0.27-0.42 Fructose  0.3-0.39Mannitol 1.6-1.9 Polysorbate 80 0.75-0.96 Tartaric Acid 0.033-0.043Sodium Hydroxide — Hydrochloric Acid — Ethanol  5-30 PEG and/orpropylene glycol  5-30 Sterile Water for Injection to volume

[0078] In a pharmaceutically acceptable aqueous parenteral formulationof an echinocandin, such as anidulafungin, the weight percent ofpolyethylene glycol or propylene glycol, if present, may be, e.g., 5 to50%; or about 20% w/v in an aqueous solvent such as water or anwater-ethanol mixture.

[0079] The molecular weight of the polyethylene glycol may be less thanabout 1500, often less than about 1000, sometimes less than about 600,and sometimes less than about 400. Polyethylene glycol having molecularweight in the range 400-600 may be optimal. In a pharmaceuticallyacceptable parenteral formulation of echinocandin, such asanidulafungin, the weight percent of polyethylene glycol may be forexample about 5 to 50% w/v or about 10 to 20% w/v in an aqueous solventcomprising ethanol.

[0080] A pharmaceutically acceptable parenteral formulation ofechinocandin such as anidulafungin can include about 10% w/v ethanol and10% w/v propylene glycol or polyethylene glycol in an aqueous solutionsuch as water. In the pharmaceutically acceptable parenteralformulations, in one embodiment the sum of the percent w/v is less thanor equal to 50.

[0081] The formulation can for example include an echinocandin such asanidulafungin at a concentration of about 0.1 to 2.0% w/v or about 0.25to 0.45% w/v or optionally 0.34% w/v.

[0082] In reconstitution of the solid composition with an aqueousethanol mixture after a long storage period of e.g. greater than oneyear, two years, three years or more, the mixing or shaking time forreconstitution may be less than about 400 seconds, often less than about300 seconds, sometimes less than about 200 seconds, sometimes less thanabout 100 seconds, and sometimes less than about 75 seconds, whereinresults may be improved in comparison to results using aqueous solventswithout ethanol.

[0083] Formulations may optionally contain a stabilizing agent. Astabilizing agent is present optionally at a concentration for example,in the range of about 0.3% to about 40% w/v, or about 1% to about 10%w/v. The term “stabilizing agent” refers to a pharmaceuticallyacceptable excipient that enhances the chemical or physical stability ofthe active ingredient in the formulation. Suitable stabilizing agentsinclude carbohydrates (e.g., as sucrose, trehalose, fructose, lactoseand mannitol), and amino acids.

[0084] Formulations may also optionally contain a solubilizing agent.The solubilizing agent is present, for example, at a concentration ofabout 0.2% to about 2.0% w/v, or about 0.75% to about 1.0% w/v. Suitablesolubilizing agents include polysorbates (eg polysorbate 20, polysorbate40, and polysorbate 80).

[0085] Formulations may also optionally contain a buffer. The buffer ispresent for example at a concentration in the range from about 0.03% toabout 5.0% w/v, or about 0.1% to about 2.0% w/v. The term “buffer”refers to a pharmaceutically acceptable excipient that maintains the pHof the solution within a particular range specific to the bufferingsystem. Formulations may have, for example, a pH of about 3.0-7.0,optionally about 4.0-6.0, or about 4.4-4.6. Suitable buffers includeacetates, citrates, phosphates, tartrates, lactates, succinates, aminoacids and the like. Sodium hydroxide and/or hydrochloric acid can beused to adjust the pH.

[0086] Formulations may optionally contain a bulking agent. The bulkingagent is for example present in a formulation at a concentration in therange from about 1% to about 60% w/v, or about 3% to about 50% w/v. Theterm “bulking agent” refers to a pharmaceutically acceptable excipientthat adds bulk to a formulation which results in a well-formed cake uponfreeze drying. Suitable bulking agents include mannitol, glycine,lactose, sucrose, trehalose, dextran, hydroxyethyl starch, ficoll andgelatin. Preferred bulking agents include mannitol, sucrose, trehalose,lactose and combinations thereof.

[0087] Formulations may further include one or more antioxidants. Theantioxidant is for example present in a formulation at a concentrationrange of 0.01-10% w/v or about 1-5% w/v. Examples of antioxidantsinclude acetone sodium bisulfite, bisulfite sodium, butylated hydroxyanisole, butylated hydroxy toluene, cystein, cysteinate HCl, dithionitesodium, gentisic acid, gentisic acid ethanolamine, glutamate monosodium,formaldehyde sulfoxylate sodium, metabisulfite potassium, metabisulfitesodium, monothioglycerol, propyl gallate, sulfite sodium, thioglycolatesodium, and ascorbic acid.

[0088] In application, the formulation is typically reconstituted andfurther diluted if necessary, prior to administration. An example ofreconstitution instructions for the lyophilized product are to addsolvent to the vial and agitated to dissolve. Typical reconstitutiontimes are less than 400 seconds. Suitable solvents include ethanol inwater. In one example, the solvent is about 20 weight percent ethanol inan aqueous solution such as water. The resulting solution is optionallyfurther diluted in an infusion solution such as dextrose 5% in water(D5W), prior to administration.

[0089] A solid composition may be reconstituted to provide a liquidproduct for parenteral administration to a patient. Solid compositionsto be reconstituted may be provided in dosage vessels which containdosage units, for example, from about 5 mg to about 500 mg ofechinocandin, such as anidulafungin, for example about 15 mg, about 25mg, about 35 mg, about 50 mg, about 100 mg, or about 200 mg. The dosagevessels are often loaded with a slight excess, for example 2.5% excessof drug, because it is generally not possible to extract all the drugupon use of the reconstituted drug. For example, a 35 mg vessel may beloaded with about 36 mg of drug. A dosage unit may be provided in asealed container, often made of Type I glass, which maintains a sterileenvironment for the solid product from lyophilization, for example, a 50ml vial hermetically sealed with a sterile rubber or plastic closure orstopper.

[0090] The solid product, for example, 35 mg of anidulafungin, andoptionally other components of the formulation, in a 10 mL vial, or 50mg anidulafungin in a 15 mL vial, can be reconstituted in apharmaceutically acceptable diluent, for example 5-50% w/v ethanol or20% w/v ethanol in Sterile Water for Injection (SWFI) to a concentrationof drug that is for example about 0.5-5 mg/mL, about 3-4 mg/ml or about3.3 mg/ml. The reconstituted drug is then further diluted about 5-10fold or about 7 fold with a pharmaceutically acceptable diluent such as5% Dextrose in water (D5W), to a concentration of, e.g., 0.1 to 3 mg/mLor about 0.5 mg/mL for administration. This solution can be administeredintravenously.

[0091] The following examples are provided to illustrate but not limitthe invention.

[0092] All documents, including publications, treatises, articles,patents and patent applications referenced herein are incorporatedherein by reference in their entirety.

EXAMPLES Example 1 Method of Manufacture of Solid AnidulafunginFormulation

[0093] Manufacture

[0094] In a suitable manufacturing vessel, 25 grams of polysorbate 80,are added to a sufficient amount of deoxygenated Sterile Water forInjection and mixed slowly until dissolved. The solution is cooled and1.1 gram of tartaric acid is added. The solution is adjusted to pH 4.5using a sodium hydroxide solution and/or a hydrochloric acid solution.

[0095] 10 grams of anidulafungin are added to a suitable vessel withSterile Water for Injection and swirled to generate a slurry. Theresulting slurry is added to the bulk polysorbate 80 buffer solution andthe liquid is mixed until all the slurried drug is dissolved. Thefructose (10 grams) and mannitol (50 grams) are added and mixed untildissolved. Additional Sterile Water for Injection is added to bring thesolution to final volume. The pH of the final solution is determined andadjusted, if necessary, to pH 4.5. The solution is sterilized bymembrane filtration using two redundant in-line 0.22μ Millipak 20filters into a Class 100 aseptic area. These membrane filters arenon-asbestos, non-fiber releasing, and meet cGMP requirements for use inthe manufacture and processing of components of drug products forparenteral injection in humans. Sterilizing filters are integrity testedafter use to assure that integrity was maintained during filtration.In-process testing occurs on the bulk material after the filtration forAppearance, Anidulafungin concentration, and pH.

[0096] Lyophilization

[0097] Aliquots (3.5 ml) of the sterile filtered bulk solution areaseptically filled into the sterile (depyrogenated) glass vials andsterile stoppers are partially inserted. The filled vials aretransferred to the freeze dryer and lyophilized.

[0098] The freeze drying process is monitored by thermocouple probes ofrepresentative vials. Vials are loaded into the freeze dryer and thetemperature is gradually reduced until all thermocouples reach −40° C.After the desired time has elapsed, the air in the chamber is evacuatedand the temperature gradually increased until the shelf temperature isapproximately +35° C. The samples are held at about +35° C. for 6-8hours. The chamber is then restored to atmospheric pressure withfiltered Nitrogen UHP and the stoppers are seated. Vials are removed andcapped with aluminum seals.

[0099] During aseptic operations such as set-up, filtration, filling andstoppering, the air is monitored for microbial content with settlingplates and volumetric air sampler. In addition, operators and surfacesare monitored by contact plates. The air is monitored for particlesusing a particle counter. Records of the results of these surveys areevaluated against pre-established action limits for the area involvedand, if necessary, an investigation is conducted. Appropriate action istaken when indicated by the results of the investigation.

[0100] All components and equipment are sterilized by appropriateprocesses. Sterilization processes uses the “overkill” approach for bothsteam and dry heat sterilization cycles. The cycle for steam autoclavingis for 30 minutes at 121° C. and the cycle for dry heat sterilization isfor at least three hours at >250° C. The resulting solid is stored atroom temperature.

[0101] Reconstitution

[0102] Reconstitution occurs in a two step process. In the first step,the solid formulation containing 35 mg anidulafungin, and optionallyother components of the formulation, is reconstituted in a 10 mlsolution of 20% w/v ethanol in Sterile Water for Injection. The mixtureis swirled by hand for 100 seconds and observed to confirm the completedissolution of the solid product in a clear bubble free solution. Thismixture is then diluted 7 fold in a solution of 5% Dextrose in SterileWater (D5W). The resulting solution is now available for IV injection.

Example 2 Reconstitution Time of Solid Anidulafungin Compositions

[0103] A: Reconstitution Time of Anidulafungin Stored for up to 36Months

[0104] Solid compositions containing 1) 35 mg anidulafungin (lotCT12759, PPD04365), 175 mg mannitol USP, 87.5 mg polysorbate 80 NF, 35mg fructose USP, and 3.95 mg tartaric acid NF as a buffer; or 2) 25 mg(lot CT12758) anidulafungin, 125 mg mannitol USP, 62.5 mg polysorbate 80NF, 25 mg fructose USP, and 2.5 mg tartaric acid NF as a buffer, weretested to determine reconstitution time after storage. The samples oflyophilized formulations, obtained from Eli Lilly (Indianapolis, Ind.),were stored in solid form for up to 36 months or longer. Storageconditions were 25° C., or 5° C. 60% relative humidity (RH). Thereconstitution time assay consisted of initiating reconstitution of thesolid composition by injecting 10 ml of solvent into the vial containing35 mg of anidulafungin and 7 ml of solvent into the vial containing 25mg of anidulafungin. Once the solvent was added, it was immediatelymixed in the vial by gently shaking or swirling by hand. Every 10seconds shaking was stopped and the vial was visually inspected untilcompleteness of solution was observed. The results in Table 1 show eachreconstitution time reported represented as an average of 5 replicates.It can be seen from the Tables that the addition of ethanol to thereconstitution mixture significantly reduces the reconstitution time ofanidulafungin.

[0105] A comparison was made between shaken and swirled (by hand)reconstitution methods. Swirling generally results in longerreconstitution times, but is used to reduce foaming. By shaking thediluent, reconstitution times are shortened, but foaming can often occurwith the use of water alone. Using aqueous ethanol as a diluent, it ispossible to use shaking to dissolve the drug quickly, and foaming isreduced.

[0106] As illustrated in Table 1, reconstitution of solid compositionscontaining anidulafungin was significantly reduced from greater than5400 seconds to 74 seconds using 20% w/w ethanol in water in comparisonwith pure water. TABLE 1 Reconstitution Times of Solid CompositionsContaining Anidulafungin Shaken in Indicated Diluent Storage StorageTime‡ Condition Lot No. Time (Seconds) diluent* Initial CT12758 Initial   24 Water 25° C./60% RH CT12758  3 month    164 Water 25° C./60% RHCT12758 18 month    338 Water 25° C./60% RH CT12758 25 month >5400 Water25° C./60% RH CT12758 36 month >5400** Water Initial PPD04365 Initial   27 Water 25° C./60% RH PPD04365  3 month    101 Water 25° C./60% RHPPD04365 18 month    271 Water 25° C./60% RH PPD04365 25 month >5400Water Initial CT12759 Initial    32 Water 25° C./60% RH CT12759  3 month   64 Water 25° C./60% RH CT12759 18 month    247 Water 25° C./60% RHCT12759 25 month >5400** Water 25° C./60% RH CT12759 36 month >5400**Water 25° C./60% RH CT12759 32 months    74 Aqueous Ethanol

[0107] Tables 2 and 3 show a comparison of reconstitution times fordifferent lots of solid anidulafungin compositions in water and aqueousethanol, where, advantageously, drug could be dissolved in 120 secondsor less with shaking using aqueous ethanol. Generally using water,shaking produces unwanted foaming. Swirling can be used to reducefoaming, but the length of reconstitution time can increaseconsiderably. Reconstitution time using water and shaking varied between150 and 360 seconds. Reconstitution times using water and swirling,varied between 240 and 1200 seconds. The reconstitution time using waterand ethanol and shaking is between 60 and 120 seconds and issignificantly reduced relative to reconstitution in water. Thus, using awater/ethanol mixture results less foaming and a reduced reconstitutiontime. TABLE 2 Reconstitution Times for Lot 12758 Stored for 3 Years at5° C. Time Lot No. Vial Diluent* Agitation (seconds) FinalReconstitution Evaluation Swirled CT12758 1 Water Swirled 1200 Hazy,with a significant number of small particles CT12758 2 Water Swirled 600Completely in solution CT12758 3 Water Swirled 1200 Clear solution witha small number of large particles CT12758 4 Water Swirled 1200 Clearsolution with a small number of small particles CT12758 5 Water Swirled720 Completely in solution Shaken CT12758 1 Water Shaken 360 Completelyin solution CT12758 2 Water Shaken 330 Completely in solution CT12758 3Water Shaken 330 Completely in solution CT12758 4 Water Shaken 360Completely in solution CT12758 5 Water Shaken 330 Completely in solutionSwirled CT12758 1 Aqueous Ethanol Swirled 300 Completely in solutionCT12758 2 Aqueous Ethanol Swirled 300 Completely in solution CT12758 3Aqueous Ethanol Swirled 300 Completely in solution CT12758 4 AqueousEthanol Swirled 300 Completely in solution CT12758 5 Aqueous EthanolSwirled 300 Completely in solution Shaken CT12758 1 Aqueous EthanolShaken 120 Completely in solution CT12758 2 Aqueous Ethanol Shaken 120Completely in solution CT12758 3 Aqueous Ethanol Shaken 120 Completelyin solution CT12758 4 Aqueous Ethanol Shaken 60 Completely in solutionCT12758 5 Aqueous Ethanol Shaken 120 Completely in solution

[0108] TABLE 3 Reconstitution Times for Lot 12759 Stored for 3 Years at5° C. Lot Vial Diluent* Agitation Time(s) Final ReconstitutionEvaluation Swirled CT12759 1 Water Swirled 480 Completely in solutionCT12759 2 Water Swirled 600 Completely in solution CT12759 3 WaterSwirled 480 Completely in solution CT12759 4 Water Swirled 720Completely in solution CT12758 5 Water Swirled 240 Completely insolution Shaken CT12759 1 Water Shaken 150 Completely in solutionCT12759 2 Water Shaken 150 Completely in solution CT12759 3 Water Shaken150 Completely in solution CT12759 4 Water Shaken 180 Completely insolution CT12758 5 Water Shaken 180 Completely in solution SwirledCT12759 1 Aqueous Ethanol Swirled 240 Completely in solution CT12759 2Aqueous Ethanol Swirled 300 Completely in solution CT12759 3 AqueousEthanol Swirled 300 Completely in solution CT12759 4 Aqueous EthanolSwirled 300 Completely in solution CT12758 5 Aqueous Ethanol Swirled 300Completely in solution Shaken CT12759 1 Aqueous Ethanol Shaken 60Completely in solution CT12759 2 Aqueous Ethanol Shaken 60 Completely insolution CT12759 3 Aqueous Ethanol Shaken 60 Completely in solutionCT12759 4 Aqueous Ethanol Shaken 90 Completely in solution CT12758 5Aqueous Ethanol Shaken 60 Completely in solution

[0109] B: Reconstitution times for Anidulafungin stored for 42 months

[0110] Solid compositions containing 1) 35 mg (lot CT12759)anidulafungin, 175 mg mannitol USP, 87.5 mg polysorbate 80 NF, 35 mgfructose USP, and 3.95 mg tartaric acid NF as a buffer; or 2) 25 mg (lotCT12758) anidulafungin, 125 mg mannitol USP, 62.5 mg polysorbate 80 NF,25 mg fructose USP, and 2.5 mg tartaric acid NF as a buffer, were testedto determine reconstitution time after storage. These samples oflyophilized formulations, obtained from Eli Lilly (Indianapolis, Ind.),were stored in solid form for 42 months. Storage conditions were 25° C.,60% relative humidity (RH). The reconstitution time assay consisted ofinitiating reconstitution of the solid composition by injecting 10 ml ofsolvent into the vial containing 35 mg of anidulafungin and 7 ml ofsolvent into the vial containing 25 mg of anidulafungin. Once thesolvent was added, it was immediately mixed in the vial by gentlyshaking or swirling by hand. Every 10 seconds shaking was stopped andthe vial was visually inspected. The assay was stopped after 1200seconds and the samples were visually evaluated for clarity andcompleteness of solution. Samples that appeared completely dissolvedwere further evaluated for particulate matter by a light obscurationassay described in USP 23 Section 788.

[0111] The results in Tables 4 and 5 show each reconstitution time for 3different vials for each lot in water that was shaken or swirled and ina 20% (w/w) ethanol in water solution that was shaken. The Tables alsoshow a description of the final reconstitution evaluation. It can beseen that the addition of ethanol to the reconstitution mixture enablesthe complete reconstitution of anidulafungin samples stored for 42months after only 200 or 300 seconds of swirling. Samples from identicallots were not completely dissolved in pure water after 1200 seconds ofshaking or swirling. The solutions resulting from reconstitution inaqueous ethanol were also significantly clearer and showed no foaming orparticulate matter in contrast to the corresponding water solutions.Measurements of particulate matter of samples reconstituted in aqueousethanol from lot CT12758 show only 95 particles greater than 10 μm insize and only 12 particles greater than 25 μm in size. Samples from lotCT12759 reconstituted in aqueous ethanol showed similarly small numbersof particles, 67 and 4 particles greater than 10 and 25 μm in sizerespectively. This significantly smaller than the USP specification of6000 and 600 particles 10 and 25 μm in size.

[0112] The samples dissolved in water were hazy and showed significantparticulate matter visible to the naked eye and therefore contained wellover the level of particulate matter to be accurately measured by thelight obscuration method. The samples reconstituted in aqueous ethanoldiluent were shaken, not swirled. This method results in significantfoaming when used with pure water as the diluent. The results wereconsistent for both of the drug lots that were evaluated. The aqueousethanol mixture acts to both reduce reconstitution time and reducefoaming of the reconstituted anidulafungin samples stored for as long as42 months. TABLE 4 Reconstitution times for Lot CT12758 (25 mg/vial)stored for 42 months at 25° C./60% RH Time (sec- Final ReconstitutionLot# Vial Diluent Agitation onds) Evaluation CT12758 1 Water* Swirled1200 Hazy, large particles observed CT12758 2 Water* Swirled 1200 Hazy,large particles observed CT12758 3 Water* Swirled 1200 Hazy, largeparticles observed CT12758 1 Water* Shaken 1200 Hazy and foamy, with asignificant number of small particles CT 12758 2 Water* Shaken 1200 Hazyand foamy, with a significant number of small particles CT12758 3 Water*Shaken 1200 Hazy and foamy, with a significant number of small particlesCT12758 1 Aq. Shaken 200 Completely in solution EtOH** CT12758 2 Aq.Shaken 200 Comletely in solution EtOH** CT12758 3 Aq. Shaken 200Completely in solution EtOH**

[0113] TABLE 5 Reconstitution times for Lot CT12759 (35 mg/vial) storedfor 42 months at 25° C./60% RH Time (sec- Final Reconstitution Lots#Vial Diluent Agitation onds) Evaluation CT12759 1 Water* Swirled 1200Clear, with a significant number of different particles CT12759 2 Water*Swirled 1200 Clear, with a significant number of different particlesCT12759 3 Water* Swirled 1200 Clear, with a significant number ofdifferent particles CT12759 1 Water* Shaken 1200 Hazy and foamy, withlarge and small particles CT12759 2 Water* Shaken 1200 Hazy and foamy,with large and small particles CT12759 3 Water* Shaken 1200 Hazy andfoamy, with large and small particles CT12759 1 Aq. Shaken 300Completely in solution EtOH** CT12759 2 Aq. Shaken 300 Completely insolution EtOH** CT12759 3 Aq. Shaken 300 Completely in solution EtOH**

Example 3 Particulate Matter, pH, and Clarity of ReconstitutedAnidulafungin

[0114] Anidulafungin solid compositions were reconstituted with 20%ethanol in water (SWFI) (w/w) or 100% water (SWFI). Solid compositionscontained 1) 35 mg anidulafungin (lot CT12759), 175 mg mannitol USP,87.5 mg polysorbate 80 NF, 35 mg fructose USP, and 3.95 mg tartaric acidNF as a buffer; or 2) 25 mg (lot CT12758) anidulafungin, 125 mg mannitolUSP, 62.5 mg polysorbate 80 NF, 25 mg fructose USP, and 2.5 mg tartaricacid NF as a buffer, and had been stored in solid form for up to 36months. The samples (from the designated Lot Nos.) were obtained fromEli Lilly (Indianapolis, Ind.) and were evaluated based on appearance ofthe reconstituted sample, pH, and particulate matter. Appearance wasvisually evaluated on the basis of color and clarity of the solutiondescribed in USP 24. The pH value was determined following the approachdescribed in USP 24 general chapter 791. Particulate matter was measuredby light obscuration as described in USP 23 Section 788. This in aninstrument based assay that measures particulate matter which istypically not detectable by visual inspection. The data show that thevisual appearance and pH of aqueous ethanol reconstituted samples, arevirtually identical to the aqueous samples, yet particulate matter issignificantly reduced. The results are shown in Table 6. Samples wereevaluated at the initial time point, and after various storage periods.The samples dissolved in the ethanol-water mixture were also evaluatedat 1, 4, 8, 24, and 48 hours following reconstitution. The samples werereconstituted with 10 ml (35 mg of anidulafungin) or 7 ml (25 mganidulafungin) of ethanol:purified water (SWFI) (20:80) or 100% water(SWFI).

[0115] The results demonstrated that particulate matter was reduced withthe aqueous ethanol formulation. Table 6 shows that reconstitution withaqueous ethanol after 32 months of storage resulted in only 255 10 μmparticles and 16 25 μm particles identified; in contrast, an average of747 10 μm and 107 25 μm particles were identified in the solutions thatwere reconstituted with pure water after only 25 months of storage.Higher numbers of particulates is undesirable for pharmaceuticaladministration, and greater than 6000 10 μm particles is not acceptablefor pharmaceutical IV use. TABLE 6 Appearance, pH, and ParticulateMatter of Reconstituted Anidulafungin Particulate Storage Matter Time(number (Time of after Appearance of particles) recon- Reconstituted 10μm/ stitution) Solution pH 25 μm Lot # Diluent 32 months Clear solution,4.6 393/121 CT12759 20:80 (Initial) EtOH:H₂O 32 months Clear solution,4.6 261/6  CT12759 20:80 (1 hour) EtOH:H₂O 32 months Clear solution, 4.6176/3  CT12759 20:80 (4 hours) EtOH:H₂O 32 months Clear solution, 4.7238/12  CT12759 20:80 (8 hours) EtOH:H₂O 32 months Clear solution, 4.7201/14  CT12759 20:80 (24 hours) EtOH:H₂O 32 months Clear solution, 4.7255/16  CT12759 20:80 (48 hours) EtOH:H₂O 25 month Clear solution, 4.23365/38  CT12759 100% H₂O (initial) 36 month sample did not 4.224874/264  CT12759 100% H₂O (initial) dissolve completely 25 month Clearsolution, 4.36 888/127 CT12758 100% H₂O (initial) 36 month Clearsolution, 4.28 6635/394  CT12758 100% H₂O (initial) 25 month Clearsolution, 4.18 988/157 PPD04365 100% H₂O (initial)

Example 4 Potency and total related substance TRS of reconstitutedAnidulafungin.

[0116] Solid compositions containing 1) 35 mg anidulafungin (lotCT12759), 175 mg mannitol USP, 87.5 mg polysorbate 80 NF, 35 mg fructoseUSP, and 3.95 mg tartaric acid NF as a buffer; or 2) 25 mg (lot CT12758)anidulafungin, 125 mg mannitol USP, 62.5 mg polysorbate 80 NF, 25 mgfructose USP, and 2.5 mg tartaric acid NF as a buffer and had beenstored in solid form for up to 36 months were measured for potency andrelated substances. Samples (from identified Lot Nos.) were obtainedfrom Eli Lilly. Potency (Pot) and TRS were determined using ahigh-pressure, liquid chromatograph (HPLC) equipped with a 15 cm×4.6 mm,3.5 micron particle size, Zorbax™ XDB-C18 column. The anidulafunginsamples were eluted with a 0.85% w/w aqueous phosphoric acid solutionand a 95% aqueous acetonitrile solution using methanol as the diluent. Agradient elution scheme was used where the ratio of the phosphoric acidsolution to the acetonitrile solution was varied from 95:5 to 59:41 to5:95 to 95:5 over a one hour period. As can be seen in Table 7, potencyand total related matter of the reconstituted anidulafungin areessentially unchanged by the addition of ethanol to the reconstitutioncomposition relative to reconstitution in 100% water. Furthermore, datawas measured for a total of 15 peaks in the HPLC chromatogram and allwere essentially unchanged in the presence of ethanol.

[0117] Table 7 below illustrates the potency and related substancesdata. It can be seen that both the water and ethanol water mixturesexhibit the same stability over time, illustrating that ethanol did notappear to interact with or decompose the drug formulation. TABLE 7Potency and Total Related Substance of Reconstituted Solid AnidulafunginFormulation Storage Total Time related -(Time substance after Potency %Area recon- relative under stitution) mg/vial % curve Lot No. Solvent 32months 36.1 103.1 3.5 CT12759 20:80 EtOH:H₂O (Initial) 32 months 36.2103.4 3.5 CT12759 20:80 EtOH:H₂O (1 hour) 32 months 36.3 103.7 3.6CT12759 20:80 EtOH:H₂O (4 hours) 32 months 36.2 103.4 3.5 CT12759 20:80EtOH:H₂O (8 hours) 32 months 36.2 103.4 3.7 CT12759 20:80 EtOH:H₂O (24hours) 32 months 36.2 103.4 3.7 CT12759 20:80 EtOH:H₂O (48 hours) 25month 35.9 102.6 4.0 CT12759 100% H₂O (initial) 36 month 36.2 103.4 3.5CT12759 100% H₂O (initial) 25 month 25.6 102.4 3.9 CT12758 100% H₂O(initial) 36 month 26.3 105.2 3.4 CT12758 100% H₂O (initial) 25 month35.0 100.0 3.9 PPD04365 100% H₂O (initial)

Example 5 Reconstituted Anidulafungin Parenteral Products

[0118] A pharmaceutically acceptable lyophilized formulation ofanidulafungin for injection is provided. For example, 35 mg or 50 mg isprovided per vial. Each vial of the 35 mg dosage form contains:anidulafungin, 35 mg; mannitol USP, 175 mg; polysorbate 80 NF, 87.5 mg;fructose USP, 35 mg; tartaric acid NF, 3.94 mg, as a buffer. Each vialof the 50 mg dosage form contains: anidulafungin, 50 mg; mannitol USP,250 mg; polysorbate 80 NF, 125 mg; fructose USP, 50 mg; tartaric acidNF, 5.63 mg, as a buffer.

[0119] An overfill of 2.5% of the formulation is provided to permitwithdrawal of the labeled amount after reconstitution. The lyophilizeddrug product is stored at controlled room temperature (15-30° C.).Unopened lyophilized vials of anidulafungin are preferably stored atcontrolled room temperature, 15-30° C. Vials are not be frozen andopened vials are not reused. Reconstituted vials or infusion bagscontaining anidulafungin are stored in the refrigerator, 2-8° C. Sincethe vials may not contain any preservative, the contents are usedimmediately after reconstitution. The infusion product is protected fromexposure to direct sunlight.

[0120] Following reconstitution with 20% alcohol USP/80% Sterile Waterfor Injection USP (w/w) (10 mL for the 35 mg anidulafungin vial, and 15mL for the 50 mg vial) anidulafungin is diluted with Dextrose Injection5% (5% Dextrose in Water, USP) prior to use, seven fold dilution. Thedrug, when diluted with Dextrose Injection 5%, is stored in arefrigerator at 2 to 8° C. and used or discarded within 24 hours.

1. A pharmaceutically acceptable parenteral formulation comprising anidulafungin and an aqueous solvent, wherein the formulation includes from about 5% w/v to about 50% w/v ethanol, and wherein the anidulafungin is stored in solid form for greater than 6 months prior to forming the formulation.
 2. The formulation of claim 1, wherein the aqueous solvent is water or saline.
 3. The formulation of claim 1, wherein the formulation includes about 10% to about 40% w/v ethanol, and about 0.2% to about 2.0% w/v anidulafungin.
 4. The formulation of claim 1, wherein the formulation includes about 15 to about 30% w/v ethanol.
 5. The formulation of claim 1, wherein the formulation includes about 20% w/v ethanol.
 6. The formulation of claim 1, further comprising about 10% to about 50% w/v of at least one of propylene glycol and polyethylene glycol.
 7. The formulation of claim 1, further comprising a stabilizing agent.
 8. The formulation of claim 7, wherein the stabilizing agent is selected from the group consisting of mannitol, histidine, lysine, glycine, sucrose, fructose, trehalose, lactose and mixtures thereof.
 9. The formulation of claim 1, further comprising a bulking agent.
 10. The formulation of claim 9, wherein the bulking agent is selected from the group consisting of mannitol, glycine, lactose, sucrose, trehalose, dextran, hydroxyethyl starch, ficoll and gelatin.
 11. The formulation of claim 1, further comprising a solubilizing agent.
 12. The formulation of claim 11, wherein the solubilizing agent is a polysorbate.
 13. The formulation of claim 11, wherein the solubilizing agent is (polysorbate 80).
 14. The formulation of claim 1, further comprising a buffer.
 15. The formulation of claim 14, wherein the buffer is selected from the group consisting of acetates, citrates, tartrates, lactates, succinates, and phosphates.
 16. The formulation of claim 1, further comprising a tonicity agent.
 17. The formulation of claim 16, wherein the tonicity agent is selected from the group consisting of glycerin, lactose, mannitol, dextrose, sodium chloride, sodium sulfate and sorbitol.
 18. The formulation of claim 1, further comprising an antioxidant.
 19. The formulation of claim 18, wherein the antioxidant is selected from the group consisting of acetone sodium bisulfite, bisulfite sodium, butylated hydroxy anisole, butylated hydroxy toluene, cysteine, cysteinate HCl, dithionite sodium, gentisic acid, gentisic acid ethanolamine, glutamate monosodium, formaldehyde sulfoxylate sodium, metabisulfite potassium, metabisulfite sodium, monothioglycerol, propyl gallate, sulfite sodium, thioglycolate sodium, and ascorbic acid.
 20. The formulation of claim 1, wherein the formulation comprises: 5.0-30% w/v ethanol; 0.1-2.0% w/v andifulafungin; 0.1-1.0% w/v of a stabilizing agent; 0.1-10.0% w/v of a bulking agent; 0.01-5.0% w/v of a buffer; and 0.1-5.0% w/v of a solubilizing agent.
 21. The formulation of claim 1, wherein the formulation comprises: 5.0-30% w/v ethanol; 0.1-2.0% w/v andifulafungin; 0.1-1.0% w/v fructose; 0.1-10.0% w/v mannitol; 0.01-5.0% w/v tartaric acid; and 0.1-5.0% w/v polysorbate
 80. 22. The formulation of claim 1, wherein the formulation comprises about 2 to 50% w/v of at least one of polyethylene glycol and propylene glycol.
 23. The formulation of claim 1, wherein the anidulafungin is stored in solid form for greater than 9 months prior to forming said formulation, and wherein said formulation is suitable for use as a parenteral formulation.
 24. The formulation of claim 1, wherein the anidulafungin is stored in solid form for greater than 12 months prior to forming said formulation, and wherein said formulation is suitable for use as a parenteral formulation.
 25. A method of preparing a parenteral pharmaceutical formulation, the method comprising combining a solid composition comprising anidulafungin and an ethanolic aqueous solvent to substantially dissolve the anidulafungin and produce an aqueous formulation including about 5% to about 50% w/v ethanol.
 26. The method of claim 25, wherein the formulation includes about 10% to about 30% w/v ethanol.
 27. The method of claim 25, wherein the aqueous solvent is water.
 28. The method of claim 25, further comprising shaking the formulation until the mixture is substantially clear.
 29. The method of claim 25, further comprising forming the solid composition comprising anidulafungin by lyophilizing an aqueous solution of anidulafungin.
 30. The method of claim 29, wherein the solid composition is formed by lyophilizing an aqueous solution of anidulafungin and an additive.
 31. The method of claim 30, wherein the additive is polyethylene glycol or propylene glycol.
 32. The method of claim 30, wherein the additive is selected from the group consisting of a stabilizing agent, a buffer, a tonicity agent and an antioxidant.
 33. The method of claim 25, wherein the solid composition comprising anidulafungin is solid composition produced for pharmaceutical use and stored for more than one year before said combining step; and wherein said combining step produces a formulation suitable for use as a pharmaceutically acceptable parenteral formulation.
 34. A kit for use in delivery of a pharmaceutically acceptable parenteral pharmaceutical formulation, the kit comprising: a first vial containing a pharmaceutically acceptable solid formulation comprising anidulafungin, wherein the anidulafungin is stored in solid form for greater than 6 months prior to reconstitution; and a second vial comprising a pharmaceutically acceptable aqueous solution of about 10 to 30% w/v ethanol.
 35. The kit of claim 34, wherein the first vial further comprises an additive selected from the group consisting of polyethylene glycol, propylene glycol, a stabilizing agent, a buffer, a tonicity agent and an antioxidant.
 36. The kit of claim 34, wherein the first vial contains a solid formulation comprising 25 to 200 mg of anidulafungin and the second vial contains 10 to 60 milliliters of aqueous solution comprising about 10 to 30% w/v ethanol.
 37. A kit for use in delivery of a pharmaceutically acceptable liquid parenteral pharmaceutical formulation, the kit comprising: a first vial containing a pharmaceutically acceptable solid formulation comprising 20 to 200 mg anidulafungin; and a second vial comprising a pharmaceutically acceptable aqueous solution of about 10 to 60 mL of an aqueous solution of about 10 to 30% w/v ethanol; wherein the solution in the second vial dissolves the solid formulation in the first vial in less than two minutes after one year of storage of the solid formulation.
 38. A kit for use in delivery of a pharmaceutically acceptable liquid parenteral pharmaceutical formulation, the kit comprising: a first vial containing a pharmaceutically acceptable solid formulation comprising 20 to 200 mg anidulafungin; and a second vial comprising a pharmaceutically acceptable aqueous solution; wherein the solution in the second vial dissolves the solid formulation in the first vial in less than two minutes after one year of storage of the solid formulation.
 39. The kit of claim 38, wherein the solution in the second vial dissolves the solid formulation in the first vial in less than two minutes after two years of storage of the solid formulation.
 40. The kit of claim 39, wherein the solution in the second vial dissolves the solid formulation in the first vial in less than two minutes after three years of storage of the solid formulation.
 41. The kit of claim 40, wherein the solution in the second vial comprises polyethylene glycol or polypropylene glycol.
 42. The kit of claim 41, wherein the solution in the second vial comprises 2-20% w/v polyethylene glycol or propylene glycol.
 43. A method of preparing a parenteral formulation, the method comprising combining a solid formulation comprising anidulafungin with a solvent in an effective amount to dissolve the anidulafungin rapidly in 200 seconds or less to produce a pharmaceutically acceptable parenteral formulation.
 44. The method of claim 43, wherein the anidulafungin is produced for pharmaceutical use and stored for more than one year prior to combining with the solvent.
 45. The method of claim 44, wherein the concentration of the anidulafungin is about 1.5 to 5 mg/mL. 